mRNA Targets for TDP-43 and FUS/TLS: Identifying Key RNA-Processing Errors in ALS

描述(申请人提供)：肌萎缩侧索硬化症(ALS)是一种成人起病的神经退行性疾病，其上下运动神经元过早丧失导致致命瘫痪，典型病程为一至五年。自2006年TDP-43被确定为ALS和额颞叶痴呆(FTLD)患者受累神经元中泛素化包涵体的主要蛋白质组分以来，我们对神经退行性变的理解出现了广泛公认的范式转变突破。事实上，现在已经确定，TDP-43是一种RNA结合蛋白，通常主要定位于细胞核，它形成细胞质聚集体，在各种不同的神经退行性疾病中改变其亚细胞定位，包括ALS、FTLD、阿尔茨海默氏症和帕金森氏病。此外，不仅在TDP-43，而且最近，在另一种RNA结合蛋白FUS/TLS中也发现了导致ALS的突变，证实了这些蛋白在ALS发病机制中的主要作用。此外，这些发现巩固了RNA加工改变对神经退化机制的关键贡献这一信息。我计划定义一组与疾病相关的TDP-43和/或FUS/TLS依赖的RNA加工改变。我建议利用现有的转基因小鼠品系以及新开发的方法，包括高通量测序和诱导多能干细胞(IPS)，以1)确定TDP-43和FUS/TLS在小鼠正常中枢神经系统RNA代谢调节中的作用，2)确定TDP-43和FUS/TLS表达导致的RNA加工变化，在转基因小鼠中携带致病突变，3)测试在小鼠、iPS细胞产生的人类运动神经元中发现的RNA加工变化，以及4)确定所观察到的与疾病相关的RNA加工变化的潜在机制，并在细胞培养和小鼠模型中测试阻断化合物对所确定的机制的有效性。 公共卫生相关性：与公共卫生相关的声明肌萎缩侧索硬化症，也被称为Lou Gehrig病，是一种毁灭性的疾病，会导致肌肉无力，在确诊后1-5年内进展为瘫痪和死亡。在美国，每年每55,000人中就有一人感染这种疾病，目前还没有有效的治疗方法。拟议的工作将有助于我们了解导致这种疾病的一连串事件，并揭示治疗干预的目标。

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder in which premature loss of upper and lower motor neurons leads to fatal paralysis with a typical disease course of one to five years. Starting from 2006 with the identification of TDP-43 as the major protein component of the ubiquitinated inclusions occurring in affected neurons of ALS and Frontotemporal Lobar Dementia (FTLD) patients, there has been what is widely recognized to be a paradigm-shifting breakthrough in our understanding of neurodegeneration. Indeed, it has now been established that TDP-43, an RNA-binding protein that is normally localized mainly in the nucleus, forms cytoplasmic aggregates, which alter its subcellular localization in a variety of different neurodegenerative conditions, including ALS, FTLD, Alzheimer's and Parkinson's disease. In addition, identification of ALS-causing mutations not only in TDP-43, but more recently, also in another RNA-binding protein, namely FUS/TLS, confirmed the primary role of these proteins in the pathogenesis of ALS. Furthermore, these discoveries solidified the message of a key contribution of RNA-processing alterations to mechanisms of neurodegeneration. I plan to define a set of disease-relevant TDP-43- and/or FUS/TLS-dependent RNA-processing alterations. I propose to utilize existing transgenic mouse lines along with newly developed methodologies including high-throughput sequencing and induced pluripotent stem (iPS) cells to 1) determine the role of TDP-43 and FUS/TLS in the regulation of RNA metabolism in the normal central nervous system in mice, 2) identify RNA-processing alterations caused by expression of TDP-43 and FUS/TLS carrying disease-causing mutations in transgenic mice, 3) test the RNA-processing changes found in mice, in human motor neurons produced from iPS cells, and 4) identify the mechanisms underlying the observed disease-related RNA-processing alterations and test the efficacy of blocking compounds on the defined mechanisms in cell culture and mouse models. PUBLIC HEALTH RELEVANCE: Statement of Relevance to Public Health Amyotrophic Lateral Sclerosis, also called 'Lou Gehrig's disease', is a devastating disease causing muscle weakness progressing to paralysis and death within 1-5 years from diagnosis. There are no effective treatments for this disease that affects 1 in approximately 55,000 people in USA every year. The proposed work will contribute to our understanding of the chain of events leading to this disease and to uncover targets for therapeutic interventions.